Pathogenic scarring and fibrosis represent important clinical problems with potentially serious consequences for patients, including impairment of normal tissue regeneration and neighboring tissue function. Our long-term goal is to identify the critical regulatory mechanisms that govern fibroblast differentiation Into myofibroblasts thereby developing novel strategies to control scarring and fibrosis. TGF- ~ is a central component of this mechanism and we recently determined that a focal adhesion protein, termed Hic-5, is induced by TGF-~ in normal human dermal fibroblasts;Hic-5 is also persistently expressed in hypertrophic scar myofibroblasts (HTSF). Importantly, Hic-5 is required for TGF-~ production in HTSF thereby regulating its own expression via the TGF-~ autocrine loop. Moreover, when Hic-5 is knocked down with RNAi the HTSF phenotype reverts to that of a normal dermal fibroblast supporting a therapeutic strategy of blocking fibrosis by targeting Hic-5 expression in myofibroblasts. We propose a two-year, ARRA sponsored, research program in which we test the hypothesis that TGF-~ induces Hic-5 expression through Rho GTPase-dependent pathway(s) thereby establishing an autocrine loop and a persistent myofibroblast phenotype.
In Aim 1, we will define the intracellular pathways that regulate TGF-~-dependent, Hic-5 expression in myofibroblasts. The mechanisms through which Hic-5 is induced in normal human fibroblasts and perpetuated in pathogenic fibroblasts (HTSFs) will be determined. Experiments will be performed using pharmacological inhibitors, genetic silencing (RNAi) and cDNA reconstitution, in vitro. The levels of Hic-5 and candidate signaling molecules will be determined and changes in activity measured using phosphorylation-specific antibodies.
In Aim 2, we will identify and analyze Hic-,5 regulated genes in hypertrophic scars. Expression (cDNA) arrays will be used to identify the genes products in HTSF that are regulated by Hic-5. These will be cross-referenced to genes that others have reported are markedly regulated in hypertrophic scars. Hic-5-dependent gene products identified from these cDNA arrays will also be ,analyzed as candidate genes that eitlier reguiate the TGF-~ autocrine loop or mediate specific myofibroblast functions, Key gene products will then be incorporated into quantitative PCR (qPCR) arrays and probed with RNA obtained from freshly resected hypertrophic scars. Immunohistology will be employed to confirm the expression and localization of these Hic-5- dependent gene products in scars. Findings obtained from this work will fill important gaps in understanding pathogenic myofibroblast regulation and suggest new therapeutic strategies to modulate fibrosis following serious burn or traumatic injury.

Public Health Relevance

In normal wound healing connective tissue fibroblasts are activated to myofibroblasts for a few days. During fibrosis, excessive myofibroblasts persist for months-years. Our goal is to elucidate the mechanisms that perpetuate these cells during fibrosis and develop novel therapies to block their longevity and functions.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM056442-14
Application #
7937774
Study Section
Surgery, Anesthesiology and Trauma Study Section (SAT)
Program Officer
Hagan, Ann A
Project Start
1997-08-01
Project End
2012-08-31
Budget Start
2010-09-01
Budget End
2012-08-31
Support Year
14
Fiscal Year
2010
Total Cost
$287,582
Indirect Cost
Name
Albany Medical College
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
190592162
City
Albany
State
NY
Country
United States
Zip Code
12208
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Dabiri, Ganary; Tumbarello, David A; Turner, Christopher E et al. (2008) TGF-beta1 slows the growth of pathogenic myofibroblasts through a mechanism requiring the focal adhesion protein, Hic-5. J Invest Dermatol 128:280-91
Dabiri, Ganary; Tumbarello, David A; Turner, Christopher E et al. (2008) Hic-5 promotes the hypertrophic scar myofibroblast phenotype by regulating the TGF-beta1 autocrine loop. J Invest Dermatol 128:2518-25
Meckmongkol, Teerin T; Harmon, Robert; McKeown-Longo, Paula et al. (2007) The fibronectin synergy site modulates TGF-beta-dependent fibroblast contraction. Biochem Biophys Res Commun 360:709-14
Dabiri, Ganary; Campaner, Anelisa; Morgan, Jeffrey R et al. (2006) A TGF-beta1-dependent autocrine loop regulates the structure of focal adhesions in hypertrophic scar fibroblasts. J Invest Dermatol 126:963-70

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